Human Leukemia Antigens: Partial Isolation and Characterization2

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Abstract
Human leukemia-associated antigens detected by nonhuman primate antisera were freed from leukemia cells by trypsin digestion, autolytic or spontaneous release, and KCI extraction; the released antigen was sedimented at 100,000×g in 4–6 hours. Electron microscope studies revealed many membrane vesicles in the ultracentrifuged pellets. Of the 3 release methods, trypsinization was most effective. Moreover, the leukemic cells after trypsin digestion were viable and yet refractory to lysis by nonhuman primate antisera to human leukemia cells. The antigenic sites were regenerated within 24 hours at either 37 or 4° C. The regeneration could be blocked by puromycin and cyclohexamide but not by actinomycin D or cytosine arabinoside. The amount of subcellular antigen released by 3.0M KCI never exceeded the amount released spontaneously by autolysis. Papain digestion solubili;zed HL-A antigens from leukemic cells but was ineffective in solubilizing leukemia-associated antigens. High yields of soluble leukemia-associated and HL-A antigens were obtained by pronase treatment. LeUkemia-associated antigens prepared by pronase digestion could not be sedimented at 60,000×g for 16 hours and were detected in the included volume of Sephadex G-200 gel filtration. The Sephadex G-200 fractions possessing the leukemia-associated antigens specific for lymphatic leukemia cells also contained HL-A12 activity. Neuraminidase treatment of leukemic cells resulted in a complete loss of leukemia-associated antigen activity from these cells. However, no antigen activity could be recovered in the supernatant after such enzyme digestion. Neuraminidase treatment of the subcellular membrane antigens isolated by tryptic digestion and the soluble antigens prepared by pronase digestion had no effect on the ability of these preparations to inhibit the leukemia antisera.

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